Cells survive in a mechanical environment of a tissue that the cells are part of. It has been shown that cell behaviors are extremely sensitive to the stiffness of the substrate on which the cells grow. For example, cell growth and movement can be guided solely by the rigidity or stiffness of the substrates: epithelial cell traction forces are proportional to substrate rigidity; matrix elasticity directs stem cell lineage specification; on microstructured anisotropic substrates, epithelial cells migrate along the direction of greatest stiffness; within a range of stiffness values spanning that of soft tissues, fibroblasts tune their internal stiffness to match that of their substrate.
Studies of interactions between cells and their surrounding environments have been receiving increased attention over the past twenty years. The results of several such studies are summarized below. Cell-substrate interactions can profoundly affect cell behavior, including adhesion, spreading, migration, division, differentiation, and internal cellular signaling. The binding interactions between cells and their substrate(s) are influenced by mechanical stimuli such as substrate stiffness and substrate curvatures. The effects of substrate stiffness on cell behaviors have been extensively studied. However, the effects of substrate curvature are not well documented. Since the substrates of cells in vivo are normally not flat, the responses of cells to substrate curvatures should also be a fundamental aspect of cell mechanosensitivity and mechanotransduction to its substrates. The importance of substrate curvature effects on cell behaviors can be illustrated by understanding the process of cell attachment and growth on curved surfaces of bones and implants in vivo.
Researchers use the term “contact guidance” to describe the orientation of cell locomotion in response to the topographic structures of the substrates. In 1976, by culturing chick heart fibroblasts on convex cylindrical glass fibers, researchers Dunn and Heath demonstrated that cells respond to substrate curvatures when the radii of the substrate curvatures are comparable to the cell sizes, and they fitted a radius of curvature of 100 μm above which the curvature effects on cell behavior were negligible.
Dunn and Ebendal showed that contact guidance on aligned collagen gels is largely a response to the three-dimensional shape of the substrate. By comparing normal and virally transformed hamster cells, Fisher and Tickle illustrated that the organization of microfilaments plays a role in determining the orientation of cells on curved surfaces.
Smeal et al., determined that substrate curvature was sufficient to influence the directional outgrowth of nerve cells by culturing nerve cells on filamentous surfaces and measuring directional growth. They found that the mean direction of neurite outgrowth aligned with the direction of minimum principal curvature, and the spatial variance in outgrowth direction was directly related to the maximum principal curvature. Maduram et al. established a dependence between cell polarity and shape by noting the presence of small molecules that alter actomyosin contractility. This finding revealed a stronger dependence on contractility for shapes having straight edges in contrast to those having curved edges.
Rumpler et al. investigated the role of substrate curvature on the growth of tissues. They reported that the local rate of the tissue formed by osteoblasts is strongly influenced by the geometrical features of the channels in an artificial three-dimensional matrix. Curvature-driven effects and mechanical forces within the tissue explained the growth patterns as demonstrated by numerical simulation and confocal laser scanning microscopy. Hwang et al. investigated the effects of microfiber diameter on the orientation of adhered cells. For this purpose, mouse fibroblast L929 cells were cultured on the surface of poly(D,L-lactic-co-glycolic acid) (PLGA) fibers of defined diameters ranging from 10 to 242 μm, and their adhesion and alignment were quantitatively analyzed. They found that the mean orientation of cells and the spatial variation of the cell alignment angle directly related to the microfiber diameter. Cells cultured on microfibrous scaffolds oriented along the long axis of the microfiber. An increase in cellular orientation along the longitudinal direction was noted as fiber diameter decreased.
Sanz-Herrera et al. proposed a cell constitutive model to mathematically simulate cell attachment on curved substrates, activated by contractile forces. They analyzed a single fiber bundle composed of microtubules and actin filaments activated by actomyosin motors. Then the model was macroscopically extended to the cytoskeletal level using homogenization.
In the above-mentioned literature, substrate curvature effects on cell behaviors were studied by experiments using glass rods or polymer fibers. To date, there is no reported experimental study on curvature effects of spherical substrates on cell behaviors, which motivated research by the present inventor and lead to the development of the described invention.